With rapid progress of molecular biology of genes, it has become very important to detect the base sequence of a specific gene. For example, detection of a gene in diagnosis of genetic disease before birth, diagnosis of cancer at molecular level or detection of a pathogen such as virus is seriously significant.
For such detection of genes, the method called hybridization has been employed (B. D. Hames and S. J. Higgins: Nucleic acid hybridization, a practical approach, IRL Press, 1985). This method is a method in which a single stranded DNA or a double stranded DNA having the base sequence complementary to a target sequence is labelled with a radioactive or non-radioactive label, then bound to the target sequence by utilizing the complementarity to the target sequence, namely hybridized with the target sequence to detect the target sequence. In this case, generally, the dot hybridization method to fix the target substance to a carrier [DNA, 4, 327-331, (1985)] or the Southern hybridization method [Molecular Cloning, p. 382, Cold Spring Harbor (1982)], etc. are practiced. However, these methods are cumbersome and require considerable amount of labors, and in spite of the efforts made for mechanical automation, it may be still impossible to analyze a large number of samples as routine works. For overcoming these problems, the hybridization method in which a probe is fixed onto a carrier has been devised [for example, T. R. Gingeras et al: Nucleic Acids Res. 15, 5373-5390], but such hybridization between liquid phase and solid phase is limited, and it cannot have become a method practically applicable with respect to sensitivity, etc. For overcoming the drawbacks of these liquid phase-solid phase hybridizations, the sandwich type liquid phase-liquid phase hybridization has been devised [for example, Ann-Christine Syvaenen et al: Nucleic Acids Res. 14, 5037-5048 (1986), Japanese Laid-Open Patent Publication No. 229068/1987]. However, these methods cannot be satisfactory with respect to high background becasue of use of a large excess of probe or sensitivity.
For improving sensitivity, a method to amplify a specific nucleic acid sequence has been developed (Japanese Laid-Open Patent Publication No. 281/1987) which corresponds to U.S. Pat. Nos. 4,683,195; 4,683,202; and 4,800,159 to Mullis et al. but the operation of hybridization is required also in this method, whereby cumbersomeness is not reduced at all.